The overall objective of this proposal is to understand a) IF-cbl, Ca++, and lipid binding structures of ileal receptor, b) ulstrastructural localization of IF-cbl receptor with and without cbl bond to it, c) to understand metabolic fate of IF during the absorption of cbl, d) to understand the effect of membrane lipids on the integration and catalytic function of IAP and Na+K+ ATPase. The antibodies obtained for ileal receptor and its isolated subunits, will be used not only for defining IF-cbl, Ca++, and lipid binding units, but also for ultrastructural localization of the ileal receptor. Sequence determination of peptides derived from ileal receptor or its subunits and IF will be carried out to understand its orientation, and any possible sequence homology that could exist between the receptor and IF. In vivo and in vitro experiments will be undertaken using [125I]-IF-57Ca]cbl to understand the metabolic fate of IF, during the uptake of [57Co]cbl by the enterocyte. Membrane bound [125I]-IF-[57Ca]cbl will be tested with extracts from various subcellular organelles to determine whether any of these fractions release cbl bound to IF, and whether any alteration in the structure of IF is responsible for the reverse. The latter possibility will be studied by comparing the properties of IF obtained from canine gastric and ileal mucosa. Membrane lipid analysis (MVM and BLM) will be undertaken in a developing rat intestine, to study whether changes in any specific or groups of lipids are responsible for the changes in the solubility and activity of enzymes noted during development. Hydrophobic anchor piece will be obtained from pure rat IAP, bound to liposomes and will be sequenced to understand the membrane orientation of IAP. The influence of membrane lipids on the kinetic behaviors of IAP and Na+K+ dependent ATPase will be studied. These enzymes will be purified from MVM (IAP) and BLM (ATPase). The studies embodied in this proposal will help us to understand the nature of the important proteins of brush borders, which are implicated in the absorption and transport functions of ileal enterocytes.